scholarly journals Importance of Basic Residues in the Nucleocapsid Sequence for Retrovirus Gag Assembly and Complementation Rescue

1998 ◽  
Vol 72 (11) ◽  
pp. 9034-9044 ◽  
Author(s):  
J. Bradford Bowzard ◽  
Robert P. Bennett ◽  
Neel K. Krishna ◽  
Sandra M. Ernst ◽  
Alan Rein ◽  
...  
Keyword(s):  
Zinc Finger ◽  
Mutational Analysis ◽  
Zinc Fingers ◽  
Leukemia Virus ◽  
Foamy Virus ◽  
Gag Proteins ◽  
Amino Terminal ◽  
C Terminus ◽  
Zinc Finger Motifs ◽  
And Function

ABSTRACT The Gag proteins of Rous sarcoma virus (RSV) and human immunodeficiency virus (HIV) contain small interaction (I) domains within their nucleocapsid (NC) sequences. These overlap the zinc finger motifs and function to provide the proper density to viral particles. There are two zinc fingers and at least two I domains within these Gag proteins. To more thoroughly characterize the important sequence features and properties of I domains, we analyzed Gag proteins that contain one or no zinc finger motifs. Chimeric proteins containing the amino-terminal half of RSV Gag and various portions of the carboxy terminus of murine leukemia virus (MLV) (containing one zinc finger) Gag had only one I domain, whereas similar chimeras with human foamy virus (HFV) (containing no zinc fingers) Gag had at least two. Mutational analysis of the MLV NC sequence and inspection of I domain sequences within the zinc-fingerless C terminus of HFV Gag suggested that clusters of basic residues, but not the zinc finger motif residues themselves, are required for the formation of particles of proper density. In support of this, a simple string of strongly basic residues was found to be able to substitute for the RSV I domains. We also explored the possibility that differences in I domains (e.g., their number) account for differences in the ability of Gag proteins to be rescued into particles when they are unable to bind to membranes. Previously published experiments have shown that such membrane-binding mutants of RSV and HIV (two I domains) can be rescued but that those of MLV (one I domain) cannot. Complementation rescue experiments with RSV-MLV chimeras now map this difference to the NC sequence of MLV. Importantly, the same RSV-MLV chimeras could be rescued by complementation when the block to budding was after, rather than before, transport to the membrane. These results suggest that MLV Gag molecules begin to interact at a much later time after synthesis than those of RSV and HIV.

SDC-3 coordinates the assembly of a dosage compensation complex on the nematode X chromosome

Development ◽  
1997 ◽  
Vol 124 (5) ◽  
pp. 1019-1031 ◽  
Author(s):  
T.L. Davis ◽  
B.J. Meyer
Keyword(s):  
Zinc Finger ◽  
X Chromosome ◽  
Dosage Compensation ◽  
Protein Complex ◽  
Terminal Region ◽  
X Chromosomes ◽  
C Elegans ◽  
Amino Terminal ◽  
Zinc Finger Motifs ◽  
Specific Association

X chromosome expression in C. elegans is controlled by a chromosome-wide regulatory process called dosage compensation that specifically reduces by half the level of transcripts made from each hermaphrodite X chromosome. This process equalizes X expression between the sexes (XX hermaphrodites and XO males), despite their two-fold difference in X chromosome dose, and thereby prevents sex-specific lethality. Dosage compensation is achieved by a protein complex that associates with X in a sex-specific fashion to modulate gene expression. SDC-3, a protein that coordinately controls both sex determination and dosage compensation, activates dosage compensation by directing the dosage compensation protein complex to the hermaphrodite X chromosomes. We show that SDC-3 coordinates this assembly through its own sex-specific association with X. SDC-3 in turn requires other members of the dosage compensation gene hierarchy for its stability and its X localization. In addition, SDC-3 requires its own zinc finger motifs and an amino-terminal region for its X association. Our experiments suggest the possible involvement of zinc finger motifs in X chromosome recognition and the amino-terminal region in interactions with other dosage compensation proteins.

scholarly journals Evolutionary Analysis of the Zinc Finger and Homeoboxes Family of Proteins Identifies Multiple Conserved Domains and a Common Early Chordate Ancestor

2020 ◽  
Vol 12 (3) ◽  
pp. 174-184
Author(s):  
Alexandra N Nail ◽  
Jeramiah J Smith ◽  
Martha L Peterson ◽  
Brett T Spear
Keyword(s):  
Zinc Finger ◽  
Cultured Cells ◽  
Zinc Fingers ◽  
Evolutionary Analysis ◽  
Vertebrate Lineage ◽  
Amino Terminal ◽  
Small Family ◽  
Sea Squirt ◽  
Wide Range ◽  
Carboxy Terminal

Abstract The Zinc Fingers and Homeoboxes (Zhx) proteins, Zhx1, Zhx2, and Zhx3, comprise a small family of proteins containing two amino-terminal C2–H2 zinc fingers and four or five carboxy-terminal homeodomains. These multiple homeodomains make Zhx proteins unusual because the majority of homeodomain-containing proteins contain a single homeodomain. Studies in cultured cells and mice suggest that Zhx proteins can function as positive or negative transcriptional regulators. Zhx2 regulates numerous hepatic genes, and all three Zhx proteins have been implicated in different cancers. Because Zhx proteins contain multiple predicted homeodomains, are associated with interesting physiological traits, and seem to be only present in the vertebrate lineage, we investigated the evolutionary history of this small family by comparing Zhx homologs from a wide range of chordates. This analysis indicates that the zinc finger motifs and homeodomains are highly similar among all Zhx proteins and also identifies additional Zhx-specific conserved regions, including a 13 amino acid amino-terminal motif that is nearly identical among all gnathostome Zhx proteins. We found single Zhx proteins in the sea lamprey (Petromyzon marinus) and in the nonvertebrate chordates sea squirt (Ciona intestinalis) and lancelet (Branchiostoma floridae); these Zhx proteins are most similar to gnathostome Zhx3. Based on our analyses, we propose that a duplication of the primordial Zhx gene gave rise to Zhx3 and the precursor to Zhx1 and Zhx2. A subsequent tandem duplication of this precursor generated Zhx1 and Zhx2 found in gnathostomes.

scholarly journals Mutational Analysis of Residues in PriA and PriC Affecting Their Ability To Interact with SSB in Escherichia coli K-12

2020 ◽  
Vol 202 (23) ◽  
Author(s):  
Anastasiia N. Klimova ◽  
Steven J. Sandler
Keyword(s):  
Escherichia Coli ◽  
Mutational Analysis ◽  
Wild Type ◽  
Content Type ◽  
Growth Defects ◽  
C Terminus ◽  
K 12 ◽  
And Function

ABSTRACT Escherichia coli PriA and PriC recognize abandoned replication forks and direct reloading of the DnaB replicative helicase onto the lagging-strand template coated with single-stranded DNA-binding protein (SSB). Both PriA and PriC have been shown by biochemical and structural studies to physically interact with the C terminus of SSB. In vitro, these interactions trigger remodeling of the SSB on ssDNA. priA341(R697A) and priC351(R155A) negated the SSB remodeling reaction in vitro. Plasmid-carried priC351(R155A) did not complement priC303::kan, and priA341(R697A) has not yet been tested for complementation. Here, we further studied the SSB-binding pockets of PriA and PriC by placing priA341(R697A), priA344(R697E), priA345(Q701E), and priC351(R155A) on the chromosome and characterizing the mutant strains. All three priA mutants behaved like the wild type. In a ΔpriB strain, the mutations caused modest increases in SOS expression, cell size, and defects in nucleoid partitioning (Par−). Overproduction of SSB partially suppressed these phenotypes for priA341(R697A) and priA344(R697E). The priC351(R155A) mutant behaved as expected: there was no phenotype in a single mutant, and there were severe growth defects when this mutation was combined with ΔpriB. Analysis of the priBC mutant revealed two populations of cells: those with wild-type phenotypes and those that were extremely filamentous and Par− and had high SOS expression. We conclude that in vivo, priC351(R155A) identified an essential residue and function for PriC, that PriA R697 and Q701 are important only in the absence of PriB, and that this region of the protein may have a complicated relationship with SSB. IMPORTANCE Escherichia coli PriA and PriC recruit the replication machinery to a collapsed replication fork after it is repaired and needs to be restarted. In vitro studies suggest that the C terminus of SSB interacts with certain residues in PriA and PriC to recruit those proteins to the repaired fork, where they help remodel it for restart. Here, we placed those mutations on the chromosome and tested the effect of mutating these residues in vivo. The priC mutation completely abolished function. The priA mutations had no effect by themselves. They did, however, display modest phenotypes in a priB-null strain. These phenotypes were partially suppressed by SSB overproduction. These studies give us further insight into the reactions needed for replication restart.

scholarly journals An unusual arrangement of 13 zinc fingers in the vertebrate gene Z13

1995 ◽  
Vol 311 (1) ◽  
pp. 219-224 ◽  
Author(s):  
T C Schulz ◽  
B Hopwood ◽  
P D Rathjen ◽  
J R Wells
Keyword(s):  
Dna Binding ◽  
Zinc Finger ◽  
Protein Interactions ◽  
Structural Organization ◽  
Zinc Fingers ◽  
Binding Activity ◽  
Protein Domain ◽  
Nucleic Acid Binding ◽  
C Terminus ◽  
Wide Range

The zinc finger is a protein domain that imparts specific nucleic acid-binding activity on a wide range of functionally important proteins. In this paper we report the molecular cloning and characterization of a novel murine zinc-finger gene, mZ13. Analysis of mZ13 cDNAs revealed that the gene expresses a 794-amino-acid protein encoded by a 2.7 kb transcript. The protein has an unusual arrangement of 13 zinc fingers into a ‘hand’ of 12 tandem fingers and a single isolated finger near the C-terminus. This structural organization is conserved with the probable chicken homologue, cZ13. mZ13 also contained an additional domain at the N-terminus which has previously been implicated in the regulation of zinc-finger transcription factor DNA-binding, via protein-protein interactions. mZ13 expression was detected in a wide range of murine embryonic and adult tissues. The structural organization of mZ13 and its expression profile suggest that it may function as a housekeeping DNA-binding protein that regulates the expression of specific genes.

STP1, a gene involved in pre-tRNA processing, encodes a nuclear protein containing zinc finger motifs

1992 ◽  
Vol 12 (6) ◽  
pp. 2633-2643
Author(s):  
S S Wang ◽  
D R Stanford ◽  
C D Silvers ◽  
A K Hopper
Keyword(s):  
Zinc Finger ◽  
Rna Splicing ◽  
Rna Binding ◽  
Zinc Fingers ◽  
Yeast Gene ◽  
Reading Frame ◽  
Trna Splicing ◽  
Intervening Sequences ◽  
Zinc Finger Motifs ◽  
Nuclear Location

STP1 is an unessential yeast gene involved in the removal of intervening sequences from some, but not all, families of intervening sequence-containing pre-tRNAs. Previously, we proposed that STP1 might encode a product that generates pre-tRNA conformations efficiently recognized by tRNA-splicing endonuclease. To test the predictions of this model, we have undertaken a molecular analysis of the STP1 gene and its products. The STP1 locus is located on chromosome IV close to at least two other genes involved in RNA splicing: PRP3 and SPP41. The STP1 open reading frame (ORF) could encode a peptide of 64,827 Da; however, inspection of putative transcriptional and translational regulatory signals and mapping of the 5' ends of mRNA provide evidence that translation of the STP1 ORF usually initiates at a second AUG to generate a protein of 58,081 Da. The STP1 ORF contains three putative zinc fingers. The first of these closely resembles both the DNA transcription factor consensus and the Xenopus laevis p43 RNA-binding protein consensus. The third motif more closely resembles the fingers found in spliceosomal proteins. Employing antisera to the endogenous STP1 protein and to STP1-LacZ fusion proteins, we show that the STP1 protein is localized to nuclei. The presence of zinc finger motifs and the nuclear location of the STP1 protein support the model that this gene product is involved directly in pre-tRNA splicing.

scholarly journals Inhibition of red blood cell development by arsenic-induced disruption of GATA-1

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Xixi Zhou ◽  
Sebastian Medina ◽  
Alicia M. Bolt ◽  
Haikun Zhang ◽  
Guanghua Wan ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Zinc Finger ◽  
Zinc Fingers ◽  
Arsenic Exposure ◽  
In Vivo Studies ◽  
Regulatory Factor ◽  
Zinc Finger Motifs ◽  
First Time

Abstract Anemia is a hematological disorder that adversely affects the health of millions of people worldwide. Although many variables influence the development and exacerbation of anemia, one major contributing factor is the impairment of erythropoiesis. Normal erythropoiesis is highly regulated by the zinc finger transcription factor GATA-1. Disruption of the zinc finger motifs in GATA-1, such as produced by germline mutations, compromises the function of this critical transcription factor and causes dyserythropoietic anemia. Herein, we utilize a combination of in vitro and in vivo studies to provide evidence that arsenic, a widespread environmental toxicant, inhibits erythropoiesis likely through replacing zinc within the zinc fingers of the critical transcription factor GATA-1. We found that arsenic interacts with the N- and C-terminal zinc finger motifs of GATA-1, causing zinc loss and inhibition of DNA and protein binding activities, leading to dyserythropoiesis and an imbalance of hematopoietic differentiation. For the first time, we show that exposures to a prevalent environmental contaminant compromises the function of a key regulatory factor in erythropoiesis, producing effects functionally similar to inherited GATA-1 mutations. These findings highlight a novel molecular mechanism by which arsenic exposure may cause anemia and provide critical insights into potential prevention and intervention for arsenic-related anemias.

scholarly journals Mutations of the Drosophila Zinc Finger-encoding Gene vielfältig Impair Mitotic Cell Divisions and Cause Improper Chromosome Segregation

2006 ◽  
Vol 17 (5) ◽  
pp. 2356-2365 ◽  
Author(s):  
Nicole Staudt ◽  
Sonja Fellert ◽  
Ho-Ryun Chung ◽  
Herbert Jäckle ◽  
Gerd Vorbrüggen
Keyword(s):  
Zinc Finger ◽  
Chromosome Segregation ◽  
Mitotic Cell ◽  
Interphase Nuclei ◽  
Cell Divisions ◽  
Zinc Finger Motifs ◽  
And Function ◽  
Functional Gain ◽  
The Brain ◽  
Developing Nervous System

We describe the molecular characterization and function of vielfältig (vfl), a X-chromosomal gene that encodes a nuclear protein with six Krüppel-like C2H2 zinc finger motifs. vfl transcripts are maternally contributed and ubiquitously distributed in eggs and preblastoderm embryos, excluding the germline precursor cells. Zygotically, vfl is expressed strongly in the developing nervous system, the brain, and in other mitotically active tissues. Vfl protein shows dynamic subcellular patterns during the cell cycle. In interphase nuclei, Vfl is associated with chromatin, whereas during mitosis, Vfl separates from chromatin and becomes distributed in a granular pattern in the nucleoplasm. Functional gain-of-function and lack-of-function studies show that vfl activity is necessary for normal mitotic cell divisions. Loss of vfl activity disrupts the pattern of mitotic waves in preblastoderm embryos, elicits asynchronous DNA replication, and causes improper chromosome segregation during mitosis.

scholarly journals Alteration of zif268 zinc-finger motifs gives rise to non-native zinc-co-ordination sites but preserves wild-type DNA recognition

1998 ◽  
Vol 333 (1) ◽  
pp. 85-90 ◽  
Author(s):  
Andrew GREEN ◽  
Bibudhendra SARKAR
Keyword(s):  
Dna Binding ◽  
Zinc Finger ◽  
Zinc Fingers ◽  
Dna Recognition ◽  
Dna Interaction ◽  
Wild Type ◽  
Zinc Finger Motifs ◽  
Eukaryotic Gene ◽  
First Time ◽  
The Relationship

Zinc fingers are among the major structural motifs found in proteins that are involved in eukaryotic gene regulation. Many of these zinc-finger domains are involved in DNA binding. This study investigated whether the zinc-co-ordinating (Cys)2(His)2 motif found in the three zinc fingers of zif268 could be replaced by a (Cys)4 motif while still preserving DNA recognition. (Cys)2(His)2-to-(Cys)4 mutations were generated in each of the three zinc fingers of zif268 individually, as well as in fingers 1 and 3, and fingers 2 and 3 together. Whereas finger 1 and finger 3 tolerate the switch, such an alteration in finger 2 renders the polypeptide incapable of DNA recognition. The protein–DNA interaction was examined in greater detail by using a methylation-interference assay. The mutant polypeptides containing the (Cys)4 motif in fingers 1 or 3 recognize DNA in a manner identical to the wild-type protein, suggesting that the (Cys)4 motif appears to give rise to a properly folded finger. Additional results indicate that a zif268 variant containing a (Cys)2(His)(Ala) arrangement in finger 1 is also capable of DNA recognition in a manner identical to the wild-type polypeptide. This appears to be the first time that such alterations, in the context of an intact DNA-binding domain, have still allowed for specific DNA recognition. Taken together, the work presented here enhances our understanding of the relationship between metal ligation and DNA-binding by zinc fingers.

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